A kind of micro- power of the blood platelet of the elastic film variable capacitance based on nanometer technique is passedSensorTechnical field
The invention belongs to technical field of medical instruments, it is related to a kind of for the new of fast qualitative detection blood platelet comprehensive functionThe micro-nano at-once monitor sensor of type.
Background technology
Blood platelet is as cell minimum in human body, except playing vital make on normal physiological haemostasisWith in pathologic thrombus formation and the role that key is similarly play in Atherosclerosis.As shown in fig. 6, blood is smallThe signal that plate is activated after skin injury, wherein：(a) when the endothelial layer on vascular wall breakage does not occur, at blood plateletIn unactivated state；(b) when endothelial layer occurs damaged, blood platelet is quickly activated, and adheres to the cell of damaged partIn epimatrix；(c) blood platelet being activated can increase surface area, the impaired vascular wall of covering by changing own form；(d)The blood platelet being activated can secrete procoagulant Factor, activate more blood platelets；(e) aggregation of the numerous blood platelets being activated looselyTampon is formed, the quick effect temporarily stopped blooding is reached；(f) blood platelet produces convergent force, makes loose tampon become to compact,So as to play the effect of thorough prolonged hemostasis.If the convergent force produced is not big enough, blood platelet is played in hemostasisEffect is extremely limited, and the tampon of relaxation easily comes off from impaired vascular wall, not only results in affected part againBleeding, can also induce other positions and embolism occur；Opposite, if blood platelet convergent force exceedes normal range (NR), in excessive livingBlood platelet under jump state can generate excessive tampon, trigger thrombus.
Increasing experiment shows that blood platelet convergent force has huge dive in terms of quantitative comprehensive assessment blood platelet work(Power.But blood platelet progress feature is detected in the world still depend on measurement blood platelet in the sound that stops blooding with research at presentThe viscoelasticity characteristic of some mid-early stage indexs and tampon during answering（Such as LTA, VerifyNow P2Y12 Assay,Platelet function Analyzer of Plateletworks, Impact-R, PFA 100 etc.）, lack and be directed to blood platelet function at the later stageThe assessment of performance, which has limited its application in scientific research and clinical diagnosis, it is difficult to which hematoblastic comprehensive function is providedConclusive strong conclusion.And the measurement to blood platelet convergent force depends on the detecting system of closed large scale at this stage, itsMeasurement accuracy is low, and can not hematoblastic mechanical interaction phenomenon under the dynamic subcellular fraction yardstick of real-time monitored.
The content of the invention
The expression of middle and later periods platelet function is accurately measured from microcosmic angle in order to realize, the present invention provides a kind ofThe blood platelet Micro-force sensor of elastic film variable capacitance based on nanometer technique.
A kind of blood platelet Micro-force sensor of the elastic film variable capacitance based on nanometer technique is that parallel-plate is variableCapacitor, including the positive plate 1 and minus plate 2 being parallel to each other, positive plate 1 is above, minus plate 2 is located at lower section, and anodeThe anode working face of plate 1 correspond to the anode working face of minus plate 2；
The positive plate 1 includes the anode body 11 of tabular, and the anode body 11 is provided with anode hole, the bottom hole of anode hole andHole wall constitutes anode material region 15 provided with anode material, the anode material region 15 by tubbiness anode golden film 31 and sunPole is drawn metering contact 32 and constituted, and the anode draws the one side that metering contact 32 is located at anode body 11；Anode body 11Another side be anode working face, anode working face be provided with silicone intermediate material layer；
The bottom surface of anode hole inner anode golden film is provided with people's fibronectin white；
The minus plate 2 includes the cathodic body 21 of tabular, and the one side of cathodic body 21 corresponding with the bottom hole of anode hole isCathodic work piece, cathodic work piece is provided with cathode material region 22, and the cathode material region 22 is by negative electrode golden film 41 and the moonDraw metering contact 42 and constitute in pole；
The material of the anode body 11 and cathodic body 21 is silicon chip；
Spacing between the anode working face and cathodic work piece is 10 microns -1000 microns；
During test, directly platelet suspension to be detected is dropped in the anode golden film 31 of tubbiness in the anode hole of positive plate 1,Anode is drawn into metering contact 32 simultaneously and negative electrode draws the electric capacity shelves that metering contact 42 connects electric impedance analyzer respectively, according to impedanceThe capacitance variation rate that analyzer is measured, converses hematoblastic coagulability.
The technical scheme further limited is as follows：
The thickness of the anode body 11 and the thickness of cathodic body 21 are 400-1000 microns.
The aperture of the anode hole is 50 microns ~ 500 microns.
The bottom surface of the anode golden film 31 of the tubbiness is the arc surface raised up.
The thickness of the anode golden film 31 and the thickness of negative electrode golden film 41 are 100nm.
The thickness of the silicone intermediate material layer 14 is 10nm, and silicone intermediate material is dimethyl siloxane（PDMS）.
The thickness of people's fibronectin white is 50nm.
10% of the area of negative electrode golden film 41 at least above the bottom area of anode golden film 31 of tubbiness.
Preparing the blood platelet Micro-force sensor of the elastic film variable capacitance based on nanometer technique includes positive plateThe big process of making two with minus plate is made, concrete operation step is as follows：
First, positive plate makes
Step（1）：Use low-pressure chemical vapor deposition（LPVCD）Method, in the two sides of anode body 11 of a piece of twin polishingThe silicon nitride thick upper 50nm of plating respectively（SiN）Silicon nitride film 12 and lower silicon nitride film 13 in film, formation；
Step（2）：Use standard ultraviolet photolithographic technology（UV Photolithography）With reaction equation ion etching technology（Reactive Ion Etching, RIE）, several a diameter of 2mm anode hole is etched on upper silicon nitride film 12,Anode body 11 is exposed；
Step（3）：The thick dimethyl-silicons of 10um are plated with spin coating hotplate technology on the lower silicon nitride film 13 of anode body 11Oxygen alkane material layer（PDMS）14；
Step（4）：Use sodium hydroxide（KOH）Wet etching technique etches away the material of anode body 11 in anode hole；
Step（5）：In order to reduce the probability of breakage of thin film applied, yields is improved, using deep reaction equation ion etching technology（DeepReactive Ion Etching, DRIE）Remaining last in the remainder and anode hole of silicon nitride film 12 in removalPartial silicon nitride 13, still, not insertion dimethyl siloxane material layer（PDMS）14；
Step（6）：By electron beam evaporation deposition technology, the golden film formation anode material of 100nm thickness is plated in anode holeRegion 15, anode material region 15 the anode golden film 31 of tubbiness and anode in anode hole are drawn metering contact 32 and constituted, whereinAnode draws the anode working face that metering contact 32 is located at anode body 11；
Step（7）：Bioactive process processing is carried out to the lower surface in the anode golden film 31 of tubbiness, people's fibronectin is coated withWhite (Human Fibrinogen) 16；Positive plate 1 is completed to make；
2nd, minus plate makes
Step（8）：Electronics evaporation coating techniques are used on the cathodic work piece of cathodic body 21, plating last layer 100nm thicknessCathode material region 22, cathode material region 22 includes negative electrode golden film 41 and negative electrode is drawn metering contact 42 and constituted；
Step（9）：One layer of photoresist sensitive to UV light is uniformly coated on cathode material region 22 with spin coating hotplate technologyLayer；
Step（10）：With standard ultraviolet photolithographic technology, the predetermined pattern on photoresist layer, on the predetermined pattern and positive plate 1Anode hole hole shape it is consistent, and spatially in correspondence with each other with aliging；
Step（11）：The gold not being photo-etched beyond the predetermined pattern that glue is protected is removed with chloroazotic acid, the He of negative electrode golden film 41 is retainedNegative electrode draws metering contact 42；
Step（12）：The photoresist that liquid will be covered in lower negative electrode golden film 41 and negative electrode extraction metering contact 42 is removed with photoresistRemove, complete minus plate 2 and make；
3rd, the assembling of blood platelet Micro-force sensor
By the dimethyl siloxane material layer of positive plate 1（PDMS）14 corresponding matchings are located at the cathode material region 22 of minus plate 2Top, that is, obtain blood platelet Micro-force sensor.
The advantageous effects of the present invention embody in the following areas：
1st, the research of micro-nano biological Micro-force sensor and blood platelet convergent force is combined by the present invention, it is intended to which fast quantification is assessedHematoblastic comprehensive function.
2nd, nowadays to platelet function detection and blood platelet physiology and pathology Journal of Sex Research are all seriously limited by experimentThe precision of detection device, the biological micro- power of the condenser type fexible film produced by micro-nano processing technology proposed by the present invention is passedSensor, can realize and carry out quantitative quickly measurement from microcosmic to blood platelet convergent force, be early detection platelet function abnormality,Blood platelet anticoagulant is instructed to develop and illustrate the formation mechenism of relevant disease there is provided necessary technical equipment and establish certainTheoretical and experiment basis.
3rd, blood platelet convergent force using to be carried out the new of comprehensive evaluating by smooth development of the invention as to platelet functionClinical criteria necessary theoretical foundation and experimental result are provided.
Brief description of the drawings
Fig. 1 is inventive sensor structural representation.
Fig. 2 is the manufacturing process schematic diagram of sensor anode.
Fig. 3 is the top view in sensor anode working face.
Fig. 4 is the top view of sensor cathode working face.
The schematic diagram of convergent force when Fig. 5 is present invention measurement coagulating platelets.
Fig. 6 is the schematic diagram that blood platelet is activated after skin injury.
Sequence number in Fig. 1-5：Positive plate 1, minus plate 2, anode body 11, upper silicon nitride film 12, lower silicon nitride film 13,Dimethyl siloxane material layer 14, anode material region 15, people's fibronectin white 16,17,18, anode golden film 31, anodeDraw metering contact 32, cathodic body 21, cathode material region 22, negative electrode golden film 41, negative electrode extraction metering contact 42.
Embodiment
With reference to embodiment, the present invention is further described.
Referring to Fig. 1, a kind of blood platelet Micro-force sensor of the elastic film variable capacitance based on nanometer technique is flatAndante variable condenser, including the positive plate 1 and minus plate 2 being parallel to each other, under positive plate 1 is above, minus plate 2 is located atSide, and the anode working face of positive plate 1 correspond to the anode working face of minus plate 2, between anode working face and cathodic work pieceSpacing be 500 microns.
Referring to Fig. 1 and Fig. 3, positive plate 1 includes the anode body 11 of tabular, and anode body 11 is provided with anode hole, anodeThe aperture in hole is 250 microns.The bottom hole and hole wall of anode hole are provided with the anode material region 15 that anode material is constituted；Anode materialMaterial region 15 is drawn metering contact 32 by the anode golden film 31 and anode of tubbiness and constituted, and the bottom surface of anode golden film 31 is to raise upArc surface, the thickness of anode golden film 31 is 100nm.Anode draws the one side that metering contact 32 is located at anode body 11；SunThe another side of pole body 11 is anode working face, and anode working face is provided with silicone intermediate material layer 14, organosiliconThe thickness of mesosome material layer 14 is 10nm, and silicone intermediate material is dimethyl siloxane（PDMS）.
The bottom surface of anode hole inner anode golden film is provided with people's fibronectin white 16, and the thickness of people's fibronectin white 16 is50nm。
Referring to Fig. 1 and Fig. 4, minus plate 2 includes the cathodic body 21 of tabular, cathodic body corresponding with the bottom hole of anode hole21 one side is cathodic work piece, and cathodic work piece is provided with cathode material region 22, and cathode material region 22 is by negative electrode goldFilm 41 and negative electrode are drawn metering contact 42 and constituted, and the thickness of negative electrode golden film 41 is 100nm, the area of negative electrode golden film 41 at least aboveThe 10% of the bottom area of anode golden film 31 of tubbiness.
The material of anode body 11 and cathodic body 21 is silicon chip, and thickness is 800 microns.
Preparing the blood platelet Micro-force sensor of the elastic film variable capacitance based on nanometer technique includes positive plateThe big process of making two with minus plate is made, concrete operation step is as follows：
First, positive plate makes
Referring to Fig. 2, the operation for making positive plate is as follows：
Step（1）：Use low-pressure chemical vapor deposition（LPVCD）Method, in the two sides of anode body 11 of a piece of twin polishingThe silicon nitride thick upper 50nm of plating respectively（SiN）Silicon nitride film 12 and lower silicon nitride film 13 in film, formation.
Step（2）：Use standard ultraviolet photolithographic technology（UV Photolithography）With reaction equation ion etching technology（Reactive Ion Etching, RIE）, several a diameter of 250 microns anodes are etched on upper silicon nitride film 12Hole, is exposed anode body 11.
Step（3）：The thick diformazans of 10um are plated with spin coating hotplate technology on the lower silicon nitride film 13 of anode body 11Radical siloxane material layer（PDMS）14.
Step（4）：Use sodium hydroxide（KOH）Wet etching technique etches away the material of anode body 11 in anode hole.
Step（5）：In order to reduce the probability of breakage of thin film applied, yields is improved, using deep reaction equation ion etching technology（Deep Reactive Ion Etching, DRIE）It is remaining in the remainder and anode hole of silicon nitride film 12 in removalLast part silicon nitride 13, still, not insertion dimethyl siloxane material layer（PDMS）14.
Step（6）：By electron beam evaporation deposition technology, the golden film formation anode of 100nm thickness is plated in anode holeMaterial area 15, anode material region 15 the anode golden film 31 of tubbiness and anode in anode hole are drawn metering contact 32 and constituted,Its Anodic draws the anode working face that metering contact 32 is located at anode body 11.
Step（7）：Bioactive process processing is carried out to the lower surface in the anode golden film 31 of tubbiness, thickness 50nm is coated withPeople's fibronectin white (Human Fibrinogen) 16；Positive plate 1 is completed to make.
2nd, minus plate makes
Step（8）：Electronics evaporation coating techniques are used on the cathodic work piece of cathodic body 21, plating last layer 100nm thicknessCathode material region 22, cathode material region 22 includes negative electrode golden film 41 and negative electrode is drawn metering contact 42 and constituted.
Step（9）：One layer of light sensitive to UV light is uniformly coated on cathode material region 22 with spin coating hotplate technologyPhotoresist layer.
Step（10）：With standard ultraviolet photolithographic technology, the predetermined pattern on photoresist layer, the predetermined pattern and positive plateThe hole shape of anode hole on 1 is consistent, and spatially in correspondence with each other with aliging.
Step（11）：The gold not being photo-etched beyond the predetermined pattern that glue is protected is removed with chloroazotic acid, negative electrode golden film is retained41 and negative electrode draw metering contact 42.
Step（12）：The light that liquid will be covered in lower negative electrode golden film 41 and negative electrode extraction metering contact 42 is removed with photoresistPhotoresist is removed, and is completed minus plate 2 and is made.
3rd, the assembling of blood platelet Micro-force sensor
By the dimethyl siloxane material layer of positive plate 1（PDMS）14 corresponding matchings are located at the cathode material region 22 of minus plate 2Top, obtains blood platelet Micro-force sensor, and be placed on the support of acrylic material.
Typical case's application of the present invention is as follows
During for detecting, used simultaneously by a pair of blood platelet Micro-force sensors, one measures to biological micro- power, anotherThen it is used as reference, influence of the experimental situation noise to biological micro-force measurement precision can be so reduced to greatest extent.EntirelyExperimentation is referring to Fig. 5, the principle of convergent force during blood platelet Micro-force sensor measurement coagulating platelets of the present invention, wherein：Fig. 5（a）For the schematic diagram of an electric capacity；It is that processing is multiple on a silicon chip due to being micro-nano processing, in practical operation micro-nano to passSensor electric capacity, Fig. 5（b）For the schematic diagram of 2 adjacent sensor capacitances on 1 silicon chip；Fig. 5（c）For platelet suspensionInstill the schematic diagram of a sensor capacitance.
During specific test, first platelet suspension is dropped in people's fibronectin white 16 of anode hole, measurement is nowCapacitance between sensor capacitance anode and negative electrode.Then coagulant is instilled on platelet suspension drop, waits platelet suspensionAfter drop solidification completely, due to hematoblastic convergent force, the dimethyl siloxane material layer 14 of anode hole bottom can be to enterprising oneStep is raised, and the negative electrode extraction metering contact 42 for now drawing metering contact 32 and minus plate 2 again by the anode of positive plate 1 is surveyedCapacitance between quantity sensor capacitor anode and negative electrode.The size changed according to the capacitive differential measured twice, can be measuredThe size of blood platelet clot retraction ability.